Protein interactions during coronavirus assembly
Protein interactions during coronavirus assembly
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DOI：
10.1128/jvi.71.12.9278-9284.1997
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发表时间：
1997-12-01
影响因子：
5.4
通讯作者：
Hogue, BG
中科院分区：
文献类型：
--
作者：
Nguyen, VP;Hogue, BG
冠状病毒在内质网和高尔基体之间的中间区室的膜上组装并获得其包膜。像其他有包膜的病毒一样，冠状病毒的组装可能依赖于蛋白质定位以及蛋白质 - 蛋白质和蛋白质 - RNA相互作用。我们以牛冠状病毒（BCV）为模型来研究病毒感染细胞中对冠状病毒组装很重要的病毒蛋白质之间的相互作用。BCV是冠状病毒的一个原型，除了刺突（S）糖蛋白、膜（M）糖蛋白和核衣壳（N）蛋白外，还表达一种额外的主要结构蛋白，即血凝素酯酶（HE）。通过共免疫沉淀在病毒感染的细胞中检测到由M、S和HE蛋白组成的复合物。动力学分析表明，S蛋白和HE在合成后各自迅速与M蛋白形成复合物，而由这三种蛋白组成的异源复合物形成得更慢。HE生物合成的动力学显示寡聚化的半衰期约为30分钟，这与由M、HE和S蛋白组成的复合物的出现相关，表明寡聚化和/或构象变化对于S - M - HE蛋白复合物的形成可能很重要。只发现HE二聚体与由这三种蛋白组成的异源复合物相关联。在HE上的寡糖链加工之前就检测到了S - M - HE蛋白复合物，表明这些蛋白复合物在糖链修剪之前的高尔基体中间区室形成。瞬时共表达和双标记免疫荧光显示HE和S蛋白与M蛋白共定位。从转染细胞中共免疫沉淀特定的HE - M和S - M蛋白复合物进一步支持了这一点，表明这些蛋白在没有其他病毒蛋白的情况下可以形成复合物。
Coronaviruses assemble and obtain their envelope at membranes of the intermediate compartment between the endoplasmic reticulum and Golgi complex. Like other enveloped viruses, coronavirus assembly is presumably dependent on protein localization and protein-protein as well as protein-RNA interactions. We have used the bovine coronavirus (BCV) as a model to study interactions between the viral proteins in virus-infected cells that are important for coronavirus assembly. BCV is a prototype for the coronaviruses that express an additional major structural protein, the hemagglutinin esterase (HE), in addition to the spike(S) glycoprotein, membrane (M) glycoprotein, and nucleocapsid (N) protein. Complexes consisting of the M, S, and HE proteins were detected in virus-infected cells by coimmunoprecipitations. Kinetic analyses demonstrated that S protein and HE each quickly formed a complex with M protein after synthesis, whereas heterocomplexes consisting of all three proteins formed more slowly. The kinetics of HE biosynthesis revealed that the half-life of oligomerization was similar to 30 min, which correlated with the appearance of complexes consisting of M, HE, and S proteins, suggesting that oligomerization and/or conformational changes may be important for the S-M-HE protein complexes to form. Only HE dimers were found associated with the heterocomplexes consisting of all three proteins. S-M-HE protein complexes were detected prior to processing of the oligosaccharide chains on HE, indicating that these protein complexes formed in a premedial Golgi compartment before trimming of sugar chains. Transient coexpressions and double-labeling immunofluorescence demonstrated that HE and S proteins colocalized with M protein. This was further supported by coimmunoprecipitation of specific HE-M and S-M protein complexes from transfected cells, indicating that these proteins can form complexes in the absence of other viral proteins.